This is a renewal application for grant R37 AI36082 Neutralization of primary HIV-1 viruses that was awarded in 2008. The overall goal is a detailed molecular understanding of how neutralizing antibodies, alone and together, inhibit HIV-1 infection of target cells, and how the antigens that they are directed to modulate the antibody response by interacting with dendritic cells and B cells. The work outlined in the proposal will be conducted in Dr. John Moore's laboratory at the Weill Comell Medical College We propose three Specific Aims: Specific Aim 1: Mechanism, stoichiometry and kinetic aspects of HIV-1 neutralization by Abs. In this Aim we will use new in vitro neutralization assays to dissect how neutralizing antibodies with different specificities block HIV-1 infection and to quantify the persistent fi-action that survives neutralization. We will use transmembrane-protein mutants to study how differential exposure of epitopes on the Env-glycoprotein complex affect the potency and extent of neutralization. Specific Aim 2: Synergy, cooperativity and breadth of HIV-1 neutralization. We will build on our finding of Env-glycoprotein heterogeneity as a source of synergy between neutralizing antibodies. Using new mathematical analyses, we will explore the relationship between synergy and cooperativity and how they affect potency and extent of neutralization of different primary HIV-1 inocula with varying degrees of heterogeneity. Specific Aim 3: Induction of immunosuppressive responses by the mannose moieties of gpl20 glycans. We will characterize how gpl20 at high concentrations that may prevail locally after immunization affect the interplay between dendritic cells and lymphocytes, including the secretion of cytokines and B-cell stimulatory factors. The pursuit of these goals will help provide in vitro correlates for which neutralizing antibodies may be most protective in vivo and inform the design of immunogens so that high titers of such antibodies can be induced.